ABSTRACT: Osteosarcomas (OS) arise more commonly during the rapid bone growth and remodeling, thus it is plausible to infer the relationship between their tumorigenesis and cell cycle disruption by genetic and epigenetic factors. Methylation data of 28 primary OS was obtained using the HM450K microarray, and compared with nine normal bone tissues. The tumor mutational burden was analyzed using data from the TruSight One sequencing panel. We identified 3,146 differentially methylated positions, and OS presented high methylation heterogeneity, with global hypomethylation and focal hypermethylation at CpG islands. There were 319 hypomethylated and 266 hypermethylated regions in tumors, located in the promoter regions of 172 and 178 genes, respectively. These genes were enriched for biological processes related to skeletal system morphogenesis, proliferation, inflammatory response and signal transduction. Six tumor suppressors presented deletions or hypermethylation (DLEC1, GJB2, HIC1, MIR149, PAX6, WNT5A), and four oncogenes presented gains or hypomethylation (ASPSCR1, NOTCH4, PRDM16, RUNX3). Most cases did not report mutations in DNMTs or TETs, but DNMT3B was recurrently gained, a possible explanation for the hypermethylation at CpG islands. Our analysis also revealed hypomethylation at 6p22, a region that contains several genes, including histones. Overall, aberrant DNA methylation was spread throughout the genome of osteosarcomas, presented with high variance, similar to other molecular alterations. While the open-sea hypomethylation likely contributes to genomic instability, enriched CpG island hypermethylation suggests the involvement of mechanisms related to silencing tumor suppressor genes in osteosarcoma tumorigenesis. These data also suggest DNMT3B probably impacting biological processes associated with early bone development.
